Curriculum in Genetics and Molecular Biology, University of North Carolina, Chapel Hill, NC 27599Integrative Program for Biological and Genome Sciences, University of North Carolina, Chapel Hill, NC 27599Departments of Biology and Genetics, University of North Carolina, Chapel Hill, NC 27599

Curriculum in Genetics and Molecular Biology, University of North Carolina, Chapel Hill, NC 27599Integrative Program for Biological and Genome Sciences, University of North Carolina, Chapel Hill, NC 27599Departments of Biology and Genetics, University of North Carolina, Chapel Hill, NC 27599

Curriculum in Genetics and Molecular Biology, University of North Carolina, Chapel Hill, NC 27599Integrative Program for Biological and Genome Sciences, University of North Carolina, Chapel Hill, NC 27599Departments of Biology and Genetics, University of North Carolina, Chapel Hill, NC 27599

Curriculum in Genetics and Molecular Biology, University of North Carolina, Chapel Hill, NC 27599Integrative Program for Biological and Genome Sciences, University of North Carolina, Chapel Hill, NC 27599Departments of Biology and Genetics, University of North Carolina, Chapel Hill, NC 27599

Abstract

Background High-resolution transcription start site (TSS) mapping in D. melanogaster embryos and cell lines has revealed a rich and detailed landscape of both cis- and trans-regulatory elements and factors. However, TSS profiling has not been investigated in an orthogonal in vivo setting. Here, we present a comprehensive dataset that links TSS dynamics with nucleosome occupancy and gene expression at unprecedented sequencing depth in the wandering third instar larva, a developmental stage characterized by large-scale shifts in transcriptional programs in preparation for metamorphosis.

Results The data recapitulate major regulatory classes of TSSs, based on peak width, promoter-proximal polymerase pausing, and cis-regulatory element density. We confirm the paucity of divergent transcription units in D. melanogaster, but also identify notable exceptions. Furthermore, we identify thousands of novel initiation events occurring at unannotated TSSs that can be classified into functional categories by their local density of histone modifications. Interestingly, a sub-class of these unannotated TSSs overlaps with functionally validated enhancer elements, consistent with a regulatory role for “enhancer RNAs” in defining transcriptional programs that are important for animal development.

Conclusions High-depth TSS mapping is a powerful strategy for identifying and characterizing low-abundance and/or low-stability RNAs. Global analysis of transcription initiation patterns in a developing organism reveals a vast number of novel initiation events that identify likely enhancer RNAs as well as other transcripts critical for animal development.

Copyright

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